It is generally believed that chromosome motion to the equator of the forming mitotic spindle requires the antagonistic action of two sister kinetochores. The only indication that a chromosome, which possesses a single kinetochore, can become positioned on the spindle equator is from studies on MUGed (Mitosis with Unreplicated Genomes) cells. However, one cannot visualize the motion of kinetochores in MUGed cells, only infer behavior from fixed cells. We used the BMIRR laser microsurgery system to create chromosome fragments possessing a single kinetochore (i.e., monocentric chromosomes) in PtK1 cells whose behavior could be clearly followed throughout mitosis. Under the appropriate conditions such monocentric chromosome fragments can be created that move to the spindle equator prior to anaphase onset. Same-cell correlative LM/3D EM studies are currently in progress to determine whether these kinetochores are attached to both poles. This work has been abstracted to the 1995 ASCB meeting.